Stroke has become a significant cause of mortality and morbidity among elderly people in industrialized countries. To address the growing burden of health care costs and the immeasurable emotional toll on families as well as affected individuals, new pharmacological agents for prevention and treatment are desperately needed. Oxygen deprivation and oxidative stress lead to release of cytochrome c, which is related in turn to the activation of Ins(1,4,5)P3 receptors (IP3R) controlling calcium release from intracellular stores. Therefore, small molecule antagonists of IP3R would be attractive therapeutic candidates to restore proper calcium signaling pathways and prevent apoptosis. To establish the feasibility of this approach, Echelon Biosciences proposes to synthesize and identify potential IP3 antagonists. Specifically, we will use in silico modeling in combination with chemical library synthesis to prepare phosphate-free IP3 mimetics that dock in the ligand binding site, as well as a series of xestospongin (a potent natural IP3R inhibitor) analogs. Candidate compounds will be screened using a medium throughput in-cell fluorescence-based assay in which the time course for calcium release by IP3 in appropriately stimulated cells is monitored in Fluo-3 plus drug-treated fibroblasts. Activity will be optimized by iterative cycles of modeling and synthesis based on biological activities observed. [unreadable] [unreadable] [unreadable]